It is the premise of this Challenge Grant that investigations of the dynamics of membrane structure and function would be tremendously facilitated by the development of robust methods for the manufacture of "suspended" bilayers that are surrounded on both sides by aqueous solutions. This application addresses Challenge Area 06-GM-104, Dynamics of membrane structure and function. Current technology consists of "supported" bilayers that are attached to a solid surface. They are inherently non-dynamic, as proteins they contain are generally immobile by nature, and an attached bilayer is not deformable. The proposed research is organized into a series of specific aims, each corresponding to different potential method(s) for generating functional supported bilayers, which we will explore in parallel, to see which one(s) meet the required criteria for a robust mezzanine platform using functional read-outs like lateral diffusion, binding reactions at the single molecule level, and membrane dynamics as exemplified by SNARE-dependent vesicle fusion. By attacking directly the single most critical technological limitation of mezzanine studies of the dynamics of membrane structure and function, and succeeding in producing planar unsupported bilayers containing functional proteins, we can meet the challenge outlined by NIGMS (Challenge Topic 06-GM-104), and jump start a great variety of insights from many laboratories in the field of membrane dynamics. This is a realistic goal within a two year frame, which will have a broad impact on membrane research, and which would not occur without the stimulus funding. PUBLIC HEALTH RELEVANCE: In the proposed project, we aim to generate freestanding (suspended) functional planar bi-layers for the study of membrane dynamics. Existing technology supported on solid surfaces, results in bi-layers where proteins are immobile and which cannot be deformed. By attacking this limitation, we can jump start a great variety of insights from many laboratories in the field of membrane dynamics.